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SUMMARY The metabolism of tetrahydrofolate (H₄PteGlu_n)‐bound one‐carbon (C₁) units (C₁metabolism) is multifaceted and required for plant growth, but it is unclear what of many possible synthesis pathways provide C₁units in specific organelles and tissues. One possible source of C₁units is via formate‐tetrahydrofolate ligase, which catalyzes the reversible ATP‐driven production of 10‐formyltetrahydrofolate (10‐formyl‐H₄PteGlu_n) from formate and tetrahydrofolate (H₄PteGlu_n). Here, we report biochemical and functional characterization of the enzyme fromArabidopsis thaliana(AtFTHFL). We show that the recombinant AtFTHFL has lowerK_mandk_catvalues with pentaglutamyl tetrahydrofolate (H₄PteGlu₅) as compared to monoglutamyl tetrahydrofolate (H₄PteGlu₁), resulting in virtually identical catalytic efficiencies for the two substrates. Stable transformation ofArabidopsisplants with the EGFP‐tagged AtFTHFL, followed with fluorescence microscopy, demonstrated cytosolic signal. Two independent T‐DNA insertion lines with impaired AtFTHFL function had shorter roots compared to the wild type plants, demonstrating the importance of this enzyme for root growth. Overexpressing AtFTHFL led to the accumulation of H₄PteGlu_n + 5,10‐methylene‐H₄PteGlu_nand serine, accompanied with the depletion of formate and glycolate, in roots of the transgenicArabidopsisplants. This metabolic adjustment supports the hypothesis that AtFTHFL feeds the cytosolic C₁network in roots with C₁units originating from glycolate, and that these units are then used mainly for biosynthesis of serine, and not as much for the biosynthesis of 5‐methyl‐H₄PteGlu_n, methionine, andS‐adenosylmethionine. This finding has implications for any future attempts to engineer one‐carbon unit‐requiring products through manipulation of the one‐carbon metabolic network in non‐photosynthetic organs. 

Abstract L-serine (Ser) and L-glycine (Gly) are critically important for the overall functioning of primary metabolism. We investigated the interaction of the phosphorylated pathway of Ser biosynthesis (PPSB) with the photorespiration-associated glycolate pathway of Ser biosynthesis (GPSB) using Arabidopsis thaliana PPSB-deficient lines, GPSB-deficient mutants, and crosses of PPSB with GPSB mutants. PPSB-deficient lines mainly showed retarded primary root growth. Mutation of the photorespiratory enzyme Ser-hydroxymethyltransferase 1 (SHMT1) in a PPSB-deficient background resumed primary root growth and induced a change in the plant metabolic pattern between roots and shoots. Grafting experiments demonstrated that metabolic changes in shoots were responsible for the changes in double mutant development. PPSB disruption led to a reduction in nitrogen (N) and sulfur (S) contents in shoots and a general transcriptional response to nutrient deficiency. Disruption of SHMT1 boosted the Gly flux out of the photorespiratory cycle, which increased the levels of the one-carbon (1C) metabolite 5,10-methylene-tetrahydrofolate and S-adenosylmethionine. Furthermore, disrupting SHMT1 reverted the transcriptional response to N and S deprivation and increased N and S contents in shoots of PPSB-deficient lines. Our work provides genetic evidence of the biological relevance of the Ser–Gly–1C metabolic network in N and S metabolism and in interorgan metabolic homeostasis.